Department of Medical Biotechnology and Translational Medicine, University of Milano, Segrate, Milano, Italy.
Glycoconj J. 2020 Dec;37(6):713-727. doi: 10.1007/s10719-020-09963-7. Epub 2020 Nov 17.
Recently, we demonstrated that the oligosaccharide portion of ganglioside GM1 is responsible, via direct interaction and activation of the TrkA pathway, for the ability of GM1 to promote neuritogenesis and to confer neuroprotection in Neuro2a mouse neuroblastoma cells. Recalling the knowledge that ganglioside GM1 modulates calcium channels activity, thus regulating the cytosolic calcium concentration necessary for neuronal functions, we investigated if the GM1-oligosaccharide would be able to overlap the GM1 properties in the regulation of calcium signaling, excluding a specific role played by the ceramide moiety inserted into the external layer of plasma membrane. We observed, by calcium imaging, that GM1-oligosaccharide administration to undifferentiated Neuro2a cells resulted in an increased calcium influx, which turned out to be mediated by the activation of TrkA receptor. The biochemical analysis demonstrated that PLCγ and PKC activation follows the TrkA stimulation by GM1-oligosaccharide, leading to the opening of calcium channels both on the plasma membrane and on intracellular storages, as confirmed by calcium imaging experiments performed with IP3 receptor inhibitor. Subsequently, we found that neurite elongation in Neuro2a cells was blocked by subtoxic administration of extracellular and intracellular calcium chelators, suggesting that the increase of intracellular calcium is responsible of GM1-oligosaccharide mediated differentiation. These results suggest that GM1-oligosaccharide is responsible for the regulation of calcium signaling and homeostasis at the base of the neuronal functions mediated by plasma membrane GM1.
最近，我们证明神经节苷脂 GM1 的寡糖部分通过直接相互作用和激活 TrkA 途径，负责 GM1 促进神经突生成和赋予 Neuro2a 小鼠神经母细胞瘤细胞神经保护的能力。回想一下神经节苷脂 GM1 调节钙通道活性的知识，从而调节神经元功能所需的细胞质钙浓度，我们研究了 GM1-寡糖是否能够在钙信号转导的调节中重叠 GM1 的特性，排除插入质膜外层的神经酰胺部分所起的特定作用。我们通过钙成像观察到，GM1-寡糖给药于未分化的 Neuro2a 细胞导致钙内流增加，这被证明是由 TrkA 受体的激活介导的。生化分析表明，GM1-寡糖通过 PLCγ 和 PKC 的激活来激活 TrkA，导致钙通道在质膜和细胞内储存库上打开，这通过用 IP3 受体抑制剂进行的钙成像实验得到证实。随后，我们发现神经突伸长在Neuro2a 细胞中被细胞外和细胞内钙螯合剂的亚毒性给药阻断，表明细胞内钙的增加是 GM1-寡糖介导的分化的原因。这些结果表明 GM1-寡糖负责调节神经元功能介导的质膜 GM1 基础上的钙信号转导和动态平衡。
